Abstract
Myostatin (MSTN) is a member of the TGF-β superfamily that negatively regulates skeletal muscle growth and differentiation. However, the mechanism by which complete MSTN deletion limits excessive proliferation of muscle cells remains unclear. In this study, we knocked out MSTN in mouse myoblast lines using a Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR/Cas9) system and sequenced the mRNA and miRNA transcriptomes. The results show that complete loss of MSTN upregulates seven miRNAs targeting an interaction network composed of 28 downregulated genes, including TGFB1, FOS and RB1. These genes are closely associated with tumorigenesis and cell proliferation. Our study suggests that complete loss of MSTN may limit excessive cell proliferation via activation of miRNAs. These data will contribute to the treatment of rhabdomyosarcoma (RMS).
Highlights
Myostatin (MSTN) [1], known as growth/differentiation factor-8 (GDF-8), is a member of the TGF-β superfamily
Rhabdomyosarcoma (RMS) [12] is a heterogeneous tumor that has been confirmed to develop as a result of genetic alterations in mesenchymal progenitor/stem cells, which express some markers of normal skeletal muscle but show an ability to proliferate indefinitely and do not completely differentiate into the muscle phenotype [13]
After the synthetic plasmid was transfected into C2C12 cells through electroporation, sequence (NLS) and Streptococcus pyogenes Cas9 nickase to observe the fluorescence of cells the cells containing MSTN sgRNA were identified with green fluorescence under a fluorescence (Figure 1A)
Summary
Myostatin (MSTN) [1], known as growth/differentiation factor-8 (GDF-8), is a member of the TGF-β superfamily. MSTN has been confirmed to be a secreted growth factor expressed predominantly in skeletal muscle [2,3] and to negatively regulate myoblast growth and differentiation [4]. MSTN controls the activation and proliferation of satellite cells, the stem cells of skeletal muscle [5]. MSTN gene mutations, e.g., in humans [6], mice [7], cattle [8] and sheep [9], result in widespread increases in skeletal muscle mass (the “double-muscled” phenotype) [10,11]. Rhabdomyosarcoma (RMS) [12] is a heterogeneous tumor that has been confirmed to develop as a result of genetic alterations in mesenchymal progenitor/stem cells, which express some markers of normal skeletal muscle but show an ability to proliferate indefinitely and do not completely differentiate into the muscle phenotype [13]. A previous study showed that MSTN is expressed and translated in the cultured RMS cell line originally derived from embryonic RMS cell lines, RD
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